THE EFFECT OF DOSE AND INTERVAL BETWEEN 5-FLUOROURACIL AND LEUCOVORINON THE FORMATION OF THYMIDYLATE SYNTHASE TERNARY COMPLEX IN HUMAN CANCER-CELLS

We examined the importance of dosing interval between leucovorin (LCV) and 5-fluorouracil (5-FU) on intracellular thymidylate synthase (TS) ternary complex, free TS and total TS protein levels in human MCF-7 br east and NCI H630 colon cancer cell lines. A 2- to 3-fold increase in total TS was noted when either cell line was exposed to 5-FU 10 mu M p lus LCV (0.01-10 mu M) compared with a 1.4- to 1.6-fold increase in to tal TS due to 5-FU 10 mu M alone. The amount of TS ternary complex for med was 2- to 3-fold higher in both cell lines treated with the combin ation of 5-FU and LCV compared with 5-FU alone. TS complex formation a nd total TS protein increased with LCV dose (0.1-10 mu M). In MCF-7 ce lls, the maximal increase in total TS protein and TS ternary complex f ormation was observed when 5-FU was delayed for 4 h after the start of LCV exposure. In NCI H630 cells, maximal total TS protein and ternary complex formation occurred when 5-FU was delayed for 18 h after the s tart of LCV exposure. The amount of free TS did not change in either c ell line whether 5-FU was given concurrently with LCV or delayed for u p to 24 h. The accumulation rate of intracellular folates in the form of higher glutamates Glu(3)-Glu(5) was rapid in MCF-7 cells (maximal f ormation after 4 h), whereas in H630 cells accumulation of higher poly glutamates continued to increase up to 18 h. The time of peak folate p olyglutamate (Glu(3)-Glu(5)) formation coincided with the lime of peak TS complex formation and total TS protein in each cell line. In these human carcinoma cell lines, the LCV dose and interval between 5-FU an d LCV play a role in increased TS total protein and TS ternary complex ; however, the amount of free TS is independent of the interval betwee n 5-FU and LCV. The time-and dose-dependent increases in TS ternary co mplex and TS total protein are associated with differences in the accu mulation of folate polyglutamates in these cell lines.